THP-1 Cells
THP-1 is a human leukemic cell line with distinct monocytic characteristics derived from the blood of a 1-year-old boy with acute monocytic leukemia (Tsuchiya Set al (1980) Int J Cancer 26:171-176). The monocytic nature of THP-1 was established using the following cytological and cytochemical criteria: 1) a-naphthyl butyrate esterase activity which could be inhibited by NaF (sodium fluoride), 2) production of lysozyme, 3) phagocytosis (the engulfing of extracellular materials) of latex particles and sensitized sheep red blood cells, and 4) ability of mitomycin C-treated THP-1 cells to activate T-lymphocytes following concanavalin A treatment. Morphologically, the cytoplasm contained small azurophilic granules, the nucleus was indented and irregularly shaped with deep folds, and the cell membrane had Fc and C3b receptors which probably function in phagocytosis.
Typical monocytes develop from monoblasts through promonocytes in the bone marrow and in their mature form have a half-life of approximately three days. Roughly 75% of the circulating monocyte pool is found along the walls of blood vessels although these cells randomly migrate into tissues and become antigen-presenting or phagocytic. Antigen-presenting monocytes include interdigitating reticular and follicular dendritic cells of the lymph nodes and skin. Phagocytic monocytes are prominent as Kupffer cells of the liver and in the lung alveoli and bone marrow.
Whereas precursor monocytes are rich in azurophilic, peroxidase-containing cytoplasmic granules, macrophages have more numerous cell surface receptors by which they monitor their environment. These include receptors for immunoglobulin, complement, growth factors, lipoproteins, peptides and polysaccharides. Binding of ligands to these receptors triggers macrophage proliferation, chemotaxis, secretion and phagocytosis.
Many human myeloid and myelomonocytic cell lines retain some ability to differentiate into more mature phenotypes in response to various internal stimuli including growth factors, lymphokines, cytokines, vitamin D derivatives, and tumor promoters and external agents such as trauma, smoking, UV irradiation, asbestos exposure, and steroids. THP-1 cells treated with the tumor promoter 12-O-tetradecanoyl-phorbol-13 acetate (TPA) are induced to stop proliferating and differentiate into macrophage-like cells which mimic native monocyte-derived macrophages both morphologically and physiologically.
These monocyte/macrophage-like cells exhibit changes in gene expression such as the coinduction of C-fos, c-jun and the down-regulation of c-myb (Auwerx, J. (1991) Experientia 47:22-31), increase in density of the complement C3b receptor, and decrease in both FcR and the adhesion molecule, CD4. In addition, THP-1 cells produce lipoprotein lipase and apolipoprotein E, associated with atherosclerotic lesions, secrete several proinflammatory cytokines, including IL-1.beta. and TNF (Cochran FR and Finch-Arietta MB (1989) Agents and Actions 27:271-273), and may elaborate powerful oxidants and tissue destroying proteases, such as the cathepsins.
A new human cysteine protease, a cathepsin C homolog (Incyte Clone 14284), has been identified among the up-regulated genes from activated THP-1 cells. It is an acidic, lysosomal dipeptidyl aminopeptidase and has 79.3% amino acid sequence identity with rat cathepsin C isolated as a rat kidney cDNA (Kominami E et al (1992) Biol Chem 373:367-73). Incyte 14284 has the conserved residues--cys at 258, his at 405, and asn at 427--of the catalytic triad and the NNS glycosylation site in the hydrophobic region which are common to the cysteine proteases.
Kominami et al (supra) reported the presence of rat cathepsin C mRNA in almost all rat tissues. Large amounts of transcript were prevalent in liver, spleen, small and large intestine, lung and kidney, moderate amounts in esophagus, stomach, and heart, and small amounts in brain, pancreas, adrenal gland and testis. Transcript prevalence appears to correlate with the expected presence and activity of monocyte/macrophages in normal tissue function.
Cultured macrophages have been used to study the processing of cathepsin C from its synthesis as a propeptide to the mature oligomeric enzyme. Both precursor and mature cathepsin C are phosphorylated and glycosylated, and it appears that oligomerization occurs prior to entry into the lysosome (Muno, D. et al (1993) Arch Biochem Biophy 306:103-10). In studies utilizing synthetic substrates, cathepsin C was shown to function as an endopeptidase in intracellular protein degradation and as an exopeptidase (dipeptidyl aminopeptidase) in cell growth and neuraminidase activation (Kuribayashi Met al (1993) J Biochem 113:441-49).
In the normal synovium, monocytes give rise to osteoclasts which are giant, multi-nuclear cells (Tezuka K-I et al. 1994 J Biol Chem 269: 1106-1109) which attach to the bone surface. These cells produce an acidic microenvironment in which minerals and organic components of the bone matrix are solubilized. Adult bone mass generally remains constant because the rates of bone deposition and absorption are equal; however, in rheumatoid arthritis, the number and activity of osteoclasts increase under the influence of growth factors and tumor promoters. Among other hydrolases, osteoclasts produce cathepsin C which plays a role in degrading collagen, laminin, elastin and other structural proteins which comprise the extracellular matrix of the bones. Once the bone is weakened, it is even more susceptible to bone resorption, tumor invasion and metastasis.
Rheumatoid arthritis is just one example of a monocyte/macrophage disorder; in others, macrophages participate in other ways. For example, in arteriosclerosis, macrophages accumulate cholesterol from blood lipoproteins and become the foam cells of human atherosclerotic lesions. Renegade activated monocytes have also been implicated in defective defense against infection, bowel damage, osteoporosis, toxic shock syndrome, and systemic lupus erythematosus.